New instrumentation and probes offer unparalleled opportunities for molecular imaging. By applying these technologies it is now possible to monitor the proximities of molecules and trace their movements and interactions in real time using multiple fluorophore combinations. At the University of New Mexico, eight NIH-funded investigators mapping brain gene expression or defining membrane trafficking, signal transduction and adhesion mechanisms form a core user group whose research depends heavily on the application of such sophisticated biological imaging technologies. A Center for Spatiotemporal Imaging has been established to develop new algorithms for three-dimensional modeling and image qunatification. Three of the investigators participating in the center and five others constitute major users of a Zeiss LSM 510 and a recently installed BioRad Radiance both located in our Shared Cancer Center Microscopy Facility. In this application we are requesting funds for the purchase of a new Zeiss scanhead equipped with META detector and 405 nm laser to be mounted on an existing Axiovert microscope. The requested insmnnentation is necessary to enable simultaneous imaging of larger numbers of fluorophores including those in the UV range. In addition, several users have secured the photoactivable GFP isoform and demonstrated the utility of this new probe for their experiments. Although the probe can be activated by a mercury lamp, as shown by our proof of principle experiments, a 405 nm laser with AOTF is essential for these applications to take advantage of the spatial and temporal imaging capabilities afforded by this probe. Half of the users routinely conduct FRET experiments and are in need of the META detector's linear spectral unmixing functions to obtain quantitative and spatial information. Close interactions with investigators at Sandia National Laboratories will allow the immediate application of new multivariate curve resolution algorithms for the quantification and analysis of the multispectral data obtained with the META detector. Thus, the user group is uniquely poised to exploit new imaging probes and technologies with the proposed instrumentation.